Cat. # BK027

Product Uses Include

• To determine whether a protein or compound alters kinesin or dynein-microtubule mobility.
• To test other proteins for microtubule mobility.
  
  

Introduction
This is a Fluorescence based motility assay.  To perform this assay effectively you will require a video microscope system with a shutter mechanism for time lapse image acquisition. Kinesin or dynein motors are coupled to the surface of a mcroscope perfusion chamber (kinesin heavy chain motor domain is included in the kit as a positive control) and fluorescent microtubules are added. The motor-induced motility of the microtubules can be monitored and measured in a fluorescence microscope.

Kit contents
The kit contains sufficient materials for 25 assays. The following components are included:

1. Tubulin, >99% pure, lyophilized (Cat. # TL238)
2. Rhodamine tubulin, lyophilized (Cat. # TL331M)
3. Kinesin heavy chain motor domain, lyophilized (positive control) (Cat. # KR01)
4. Antifade
5. ATP (Cat. # BSA04)
6. Microtubule polymerization buffer
7. Tubulin glycerol buffer
8. Microtubule resuspension buffer
9. Blocking solution
10. Chamber wash buffer
11. Motility buffer
12. Paclitaxel (Cat. # TXD01)
13. DMSO
14. 25 acid washed perfusion chambers
15. Chamber filters
16. Manual with detailed protocols and extensive troubleshooting guide
    
    

Equipment needed

1. Fluorescence microscope equipped with filters to detect rhodamine and ready for time-lapse image capturing
   
   

Example results
The motility of microtubules in a kinesin heavy chain (Cat. # KR01) coated chamber was followed with time lapse microscopy (Fig. 1)

Figure 1. Microtubule Motility Assay. The field was selected to show a sparse area of MT coverage in order to allow clear visualization of MT motility. Each frame represents a 5 min time-point, from 0 to 15 min. The position of each MT at time zero is marked by an arrow. The arrow position remains identical in each frame to serve as a reference point for MT movement. To obtain an average MT velocity, we recommend measure motility of approximately 30 - 50 individual MTs.

Examples of publications where this product was used
Abaza, A., Soleilhac, J. M., Westendorf, J., Piel, M., Crevel, I., Roux, A. and Pirollet, F. (2003). M phase phosphoprotein 1 is a human plus-end-directed kinesin-related protein required for cytokinesis. J. Biol. Chem. 278, 27844-27852.